Differential effects of microRNAs miR-21, miR-99 and miR-145 on lung regeneration and inflammation during recovery from influenza pneumonia.

In a mouse model of influenza pneumonia, we previously documented that proliferating alveolar type II (AT2) cells are the major stem cells involved in early lung recovery. Profiling of microRNAs revealed significant dysregulation of specific ones, including miR-21 and miR-99a. Moreover, miR-145 is known to exhibit antagonism to miR-21. This follow-up study investigated the roles of microRNAs miR-21, miR-99a, and miR-145 in the murine pulmonary regenerative process and inflammation during influenza pneumonia. Inhibition of miR-21 resulted in severe morbidity, and in significantly decreased proliferating AT2 cells due to impaired transition from innate to adaptive immune responses. Knockdown of miR-99a culminated in moderate morbidity, with a significant increase in proliferating AT2 cells that may be linked to PTEN downregulation. In contrast, miR-145 antagonism did not impact morbidity nor the proliferating AT2 cell population, and was associated with downregulation of TNF-alpha, IL1-beta, YM1, and LY6G. Hence, a complex interplay exists between expression of specific miRNAs, lung regeneration, and inflammation during recovery from influenza pneumonia. Inhibition of miR-21 and miR-99a (but not miR-145) can lead to deleterious cellular and molecular effects on pulmonary repair and inflammatory processes during influenza pneumonia.

Insights into Early Recovery from Influenza Pneumonia by Spatial and Temporal Quantification of Putative Lung Regenerating Cells and by Lung Proteomics.

During influenza pneumonia, the alveolar epithelial cells of the lungs are targeted by the influenza virus. The distal airway stem cells (DASCs) and proliferating alveolar type II (AT2) cells are reported to be putative lung repair cells. However, their relative spatial and temporal distribution is still unknown during influenza-induced acute lung injury. Here, we investigated the distribution of these cells, and concurrently performed global proteomic analysis of the infected lungs to elucidate and link the cellular and molecular events during influenza pneumonia recovery. BALB/c mice were infected with a sub-lethal dose of influenza H1N1 virus. From 5 to 25 days post-infection (dpi), mouse lungs were subjected to histopathologic and immunofluorescence analysis to probe for global distribution of lung repair cells (using P63 and KRT5 markers for DASCs; SPC and PCNA markers for AT2 cells). At 7 and 15 dpi, infected mouse lungs were also subjected to protein mass spectrometry for relative protein quantification. DASCs appeared only in the damaged area of the lung from 7 dpi onwards, reaching a peak at 21 dpi, and persisted until 25 dpi. However, no differentiation of DASCs to AT2 cells was observed by 25 dpi. In contrast, AT2 cells began proliferating from 7 dpi to replenish their population, especially within the boundary area between damaged and undamaged areas of the infected lungs. Mass spectrometry and gene ontology analysis revealed prominent innate immune responses at 7 dpi, which shifted towards adaptive immune responses by 15 dpi. Hence, proliferating AT2 cells but not DASCs contribute to AT2 cell regeneration following transition from innate to adaptive immune responses during the early phase of recovery from influenza pneumonia up to 25 dpi.